New Quality-Control Investigations on Vaccines

13 Pages·2017·0.98 MB·English

International Journal of Vaccines & Vaccination
Research Article
Open Access
New quality-control investigations on vaccines:
micro- and nanocontamination
Abstract Volume 4 Issue 1 - 2017
Vaccines are being under investigation for the possible side effects they can cause. In order Antonietta M Gatti,1,2 Stefano Montanari3
to supply new information, an electron-microscopy investigation method was applied to
1National Council of Research of Italy, Institute for the Science
the study of vaccines, aimed at verifying the presence of solid contaminants by means of
and Technology of Ceramics, Italy
an Environmental Scanning Electron Microscope equipped with an X-ray microprobe. The
2International Clean Water Institute, USA
results of this new investigation show the presence of micro- and nanosized particulate 3Nanodiagnostics srl, Italy
matter composed of inorganic elements in vaccines’ samples which is not declared among the
components and whose unduly presence is, for the time being, inexplicable. A considerable Correspondence: Dr. Antonietta Gatti, National Council of
part of those particulate contaminants have already been verified in other matrices and Research of Italy, c/o Nanodiagnostics Via E. Fermi, 1/L, 41057
reported in literature as non biodegradable and non biocompatible. The evidence collected San Vito (MO), Italy, Tel [phone],
is suggestive of some hypotheses correlated to diseases that are mentioned and briefly Email
discussed.
Received: November 30, 2016 | Published: January 23, 2017
Keywords: vaccine, disease, contamination, protein corona, biocompatibility, toxicity,
nanoparticles, immunogenicity, foreign body, environment, industrial process, quality
control
Introduction was administered. For instance, Complex Regional Pain Syndrome
(CRPS), Postural Orthostatic Tachycardia Syndrome (POTS), and
Vaccines are one of the most notable inventions meant to protect Chronic Fatigue Syndrome (CFS).13 The side-effects that can arise
people from infectious diseases. The practice of variolation is century- within a relatively short time can be local or systemic.
old and is mentioned in Chinese and Indian documents dated around
1000 A.D. Over time, variolation has been replaced by vaccination, Pain at the site of injection, swelling and uncontrollable movement
vaccines have been enhanced as to technology, and the vaccination of the hands (though this last symptom can also be considered
practice is now standardized worldwide. systemic) are described. Among the systemic effects, fever, headache,
irritability, epileptic seizures, temporary speech loss, lower limbs
Side effects have always been reported but in the latest years it dysaesthesia and paresis, hot flashes, sleep disorders, hypersensitivity
seems that they have increased in number and seriousness, particularly reactions, muscle pain, recurrent syncope, constant hunger, significant
in children as the American Academy of pediatrics reports.1,2 For gait impairment, incapacity to maintain the orthostatic posture are
instance, the diphtheria-tetanus-pertussis (DTaP) vaccine was linked reported.14
to cases of sudden infant death syndrome (SIDS);3 measles-mumps-
rubella vaccine with autism;4,5 multiple immunizations with immune It is a matter of fact that every day millions of vaccine doses are
disorders;6 hepatitis B vaccines with multiple sclerosis, etc. administered and nothing notable happens, but it is also irrefutable
that, regardless of the amount of side effects that are not recorded
The notice of Tripedia DTaP by Sanofi Pasteur reports “Adverse and the percentage of which remains in fact unknown, in a limited
events reported during post-approval use of Tripedia vaccine include number cases something wrong occurs. No satisfactory explanation
idiopathic thrombocytopenic purpura, SIDS, anaphylactic reaction, or, in many cases, no explanation at all has been given and it seems
cellulitis, autism, convulsion/grand mal convulsion, encephalopathia, that those adverse effects happen on a random and stochastic basis.
hypotonia, neuropathy, somnolence and apnea”. The epidemiological
studies carried out did not show a clear evidence of those associations, Those situations induced us to verify the safety of vaccines from
even if in 2011 the National Academy of Medicine (formerly, IOM) a point of view which was never adopted before: not a biological, but
admitted: “Vaccines are not free from side effects, or adverse effects”.7 a physical approach. So, we developed a new analysis method based
on the use of a Field Emission Gun Environmental Scanning Electron
Specific researches on components of the vaccines like adjuvants Microscope investigations to detect possible physical contamination
(in most instances, Aluminum salts) are already indicated as possible in those products.
responsible of neurological symptoms8‒10 and in some cases, in-vivo
tests and epidemiological studies demonstrated a possible correlation Materials and methods
with neurological diseases.10,11 Neurological damages induced in
44 types of vaccines coming from 2 countries (Italy and France)
patients under hemodialysis treated with water containing Aluminum
were analyzed. Table 1 groups them in terms of name, brand and
are reported in literature.12
purpose.
Recently, with the worldwide-adopted vaccines against Human
Some vaccines, in fact a minority, are meant to deal with a single
Papillomavirus (HPV), the debate was reawaken due to some adverse
bacterium or virus, while others are multi-valent. The list of vaccines
effects reported by some young subjects.
we analyzed may contain repeated names, because we considered
Specific studies communicated the existence of symptoms related different batches and years of production of the same vaccine: the
to never-described-before syndromes developed after the vaccine ones against influenza in particular.
Submit Manuscript Int J Vaccines Vaccin. 2017;4(1):7‒14. 7
| http://medcraveonline.com ©2017 Gatti et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which
permits unrestrited use, distribution, and build upon your work non-commercially.
Copyright:
New quality-control investigations on vaccines: micro- and nanocontamination ©2017 Gatti et al. 8
Table 1 List of vaccines analyzed, according to their purpose
Brand Name, Country Production Batch,
N Name Description
of Distribution Expiry Date
1 Vivotif Berna Berna Biotech SA, Italy Anti-Thyphoid Vaccine (Live), group Ty21a 3000336 [2004]
2 Typhim Vi Aventis Pasteur MSD, Italy Anti-Salmonella typhi Vaccine U1510-2 [2004]
3 Typherix GlaxoSmithKline S.p.a., Italy Anti-Thypoid Vaccine (polysaccharide Vi) ATYPB061BB [2009]
4 Anatetall Chiron (now Novartis) Italy Adsorbed anti-Tetanus Vaccine 030106 [2004]
Novartis Vaccines and
5 Anatetall Adsorbed anti-Tetanus Vaccine 060510 [2009]
Diagnostics, Italy
6 Tetabulin Baxter AG, Italy Adsorbed anti-Tetanus Vaccine VNG2G006A [2009]
Novartis Vaccines and
7 Dif-Tet-All Adsorbed anti-Tetanus and diphtheria Vaccine 070501 [2009]
Diagnostics, Italy
8 Infanrix GlaxoSmithKline S.p.a., Italy Anti-Diphtheria, tetanus and pertussis vaccine AC14B071AJ [2009]
GlaxoSmithKline Biologicals Anti-diphtheria, tetanus, pertussis, hepatitis B, poliomyelitis
9 Infanrix hexa A21CC512A [ 2017]
s, Italy and disease caused by Haemophilus influenzae type b
GlaxoSmithKline Biologicals Anti-diphtheria, tetanus, pertussis, hepatitis B, poliomyelitis
10 Infanrix hexa A21CC421A [ 2017]
s. a. France and disease caused by Haemophilus influenzae type b
M-M-R vaxPro (measles, mumps, and rubella) analyzed in
11 M-M-R vaxPro Sanofi Pasteur MSD, Italy L012437 [ 2017]
Cambridge
12 Repevax Sanofi Pasteur MSD, France Anti-diphtheria-tetanus-pertussis-polio-vaccine L0362-1 [2017]
Sanofi Pasteur MSD SNC
13 Repevax Anti-diphtheria-tetanus-pertussis-polio-vaccine L0033-1 [2016]
France
14 Priorix GlaxoSmithKline S.p.a., Italy Anti--measles-mumps, and rubella (MMR) vaccine A69CB550A [2009]
Chiron (now Novartis, ),
15 Morupar Anti-measles- mumps, and rubella (MMR) vaccine 7601 [2004]
Italy
16 Varilrix GlaxoSmithKline S.p.a., Italy Anti-Chicken pox vaccine (group OKA) A70CA567A [2009]
17 Stamaril Pasteur Sanofi Pasteur MSD, Italy anti-yellow fever vaccine A5329-6 [2009]
Allergoid-Adsorbat
18 Allergopharma, Germany Antiallergic vaccine Ch-B.:30005999-B [2006]
6-Graser Starke B.
19 Engerix-B GlaxoSmithKline S.p.a., Italy Adsorbed anti-hepatitis B vaccine AHBVB468BD [2009]
20 Prenevar 13 Pfizer, Italy Antipneumococcal vaccine G79324 [2013]
21 Prevenar 13 Pfizer, France Antipneumococcal vaccine N27430 [ 2018]
anti-Neisseria meningococcal group A, C, W135 and Y
22 Mencevax Acwy GlaxoSmithKline, Italy N402A47B 12 [2004]
vaccine
23 Meningitec Pfizer, Italy (group C 10) (adsorbed on Al-Phosphate) H92709 [2015]
Anti-meningococcus (group C 10) vaccine (adsorbed on
24 Meningitec Pfizer-Italy H20500 [2014]
Al-Phosphate)
Anti-meningococcus vaccine sequestred by Procura della
25 Meningitec Pfizer-Italy G76673 [2014]
Repubblica
Anti-meningococcus vaccine sequestred by Procura della
26 Meningitec Pfizer-Italy H99459 [2015]
Repubblica
Anti-meningococcus vaccine sequestred by Procura della
27 Meningitec Pfizer-Italy H52269 [2015]
Repubblica
Novartis Vaccines and
28 Menjugate Anti-meningococcus group C YA0163AB [2010]
Diagnostics
Novartis Vaccines and
29 Menveo Antimeningococcus groups A, C, W135, Y A15083 [2017]
Diagnostics
Wyeth Pharmaceutical -
30 Meningitec Anti-meningococcus group C vaccine E83920 [2011]
France
31 Inflexal V Berna Biotech Anti-flu vaccine 2008/2009 3001463-01 [2009]
32 Vaxigrip Sanofi Pasteur MSD Anti-flu vaccine 2008/2009 D9703-1 [2009]
33 Vaxigrip Sanofi Pasteur Anti-flu vaccine 2012/2013 J8401-1 [2013]
34 Vaxigrip Sanofi Pasteur, Italy Anti-flu vaccine, with inactivated and split virus M7319-1 [2016]
Novartis Vaccines and
35 Focetria Anti-pandemic flu H1N1 vaccine 0902401 [2010]
Diagnostics
36 Agrippal Novartis Anti-flu vaccine 2012/2013 127002A [2013]
37 Agrippal Novartis vaccines, Italy Anti-flu vaccine with inactivated and split virus 2015/2016 - 152803 [2016]
Novartis Vaccines and
38 Agrippal S1 Anti-flu inactivated/superficial antigene v - 2014/2015 147302A [2015]
Diagnostics
39 Fluarix GlaxoSmithKline - GSK Anti-flu vaccine 2013 AFLUA789AA [2014]
Novartis Vaccines and
40 Fluad Anti-flu inactivated/superficial antigene vaccine - 2014/2015 142502 [2015]
Diagnostics
Citation: Gatti AM, Montanari S. New quality-control investigations on vaccines: micro- and nanocontamination. Int J Vaccines Vaccin. 2017;4(1):7‒14.
DOI: 10.15406/ijvv.2017.04.00072
Copyright:
New quality-control investigations on vaccines: micro- and nanocontamination ©2017 Gatti et al. 9
Table Continued...
Brand Name, Country Production Batch,
N Name Description
of Distribution Expiry Date
41 Gardasil Sanofi Pasteur MSD, Italy Anti-HPV types 6,11,16,18 vaccine NP01250 [2012]
42 Gardasil Sanofi Pasteur MSD, Italy Anti-HPV (types 6,11,16,18) vaccine K023804 [2016]
GlaxoSmithKline Biological,
43 Cervarix Anti-HPV (type 16,18) AHPVA238AX [2017]
Italy
anti-panleucopenia, infectious rhinotracheitis and infections
44 Feligen CRP Virbac S.A. - Carros - Italy 3R4R [2013]
by Calcivirus, veterinary Vaccine for cats
Table 2 List of the vaccines according to their manufacturers with the chemical composition of the debris identified in each sample. The elements most
represented are reported
N Company Name Alluminum Elements Identified
1 Allergopharma - Germany Allergoid yes Al
2 Aventis Pasteur MSD Lyon - Francie Typhim Vi no BrKP, PbSi, FeCr, PbClSiTi
3 Baxter AG Tetabulin no SiMg, Fe, SiTiAl, SBa, Zn
4 Berna Biotech Vivotif Berna no FeAl, ZrAlHf, SrAl, BiAlCl
5 Berna Biotech Inflexal V no CuSnPbZn, Fe, CaSiAl, SiAl, NaPZn, ZnP, AlSiTi
6 Chiron Anatetall Al(OH)3 FeAl, SZnBaAl
7 Chiron Morupar no /
8 GlaxoSmithKline- Belgium Mencevax ACWY no FeCrNi, ZrAl, FeCrNiZrAlSi
9 GlaxoSmithKline Infanrix Al(OH)3 Al, AlTi, AlSi
SBa, FeCu, SiAl, FeSi, CaMgSi, AlCaSi, Ti, Au, SCa,
10 GlaxoSmithKline Biologicals Infanrix hexa Al(OH)3
SiAlFeSnCuCrZn, CaAlSi
11 GlaxoSmithKline Biologicals Infanrix hexa Al(OH)3 ,AlPO4. 2H2O W, FeCrNi, Ti
12 GlaxoSmithKline Typherix no Ti, TiW, AlSiTiWCr, SBa, W, SiAl, AlSiTi
13 GlaxoSmithKline Priorix no WCa, WFeCu, SiAl, SiMg, PbFe, Ti, WNiFe
14 GlaxoSmithKline Engerix-B no Al (precipitates)
15 GlaxoSmithKline Varilrix no FeZn, FeSi, AlSiFe, SiAlTiFe, MgSi, Ti, Zr, Bi
AlCu, Fe, AlBi, Si, SiZn, AlCuFe, SiMg, SBa, AlCuBi, FeCrNi,
16 GlaxoSmithKline Fluarix no
SPZn
17 GlaxoSmithKline Biologicals Cervarix Al(OH)3 AlSi,FeAl, SiMg,CaSiAl, CaZn, FeAlSi, FeCr, CuSnPb
18 Novartis Vaccines and Diagnostics Anatetall Al(OH)3 Al, FeCrNi, AlCr, AlFe, BaS, ZnAl
19 Novartis Vaccines and Diagnostics Dif-Tet-All Al(OH)3 Fe,SBa, SiSBa, AlZnCu, AlZnFeCr
20 Novartis Vaccines and Diagnostics Menjugate kit Al(OH)3 SiAl,Ti,FeZn, Fe, Sb, SiAlFeTi, W, Zr
Fe, FeCrNiCu, FeCrNi, SiFeCrNi, Cr, SiAlFe, AlSiTiFe, AlSi,
21 Novartis Vaccines and Diagnostics Focetria no
SiMgFe, Si, FeZn
Ca, Fe, SBa, SBaZn, Cr, Si, Pb, Bi, e FeSiAlCr, SiAlSBaFe, CaAlSi,
22 Novartis Agrippal S1 no
Zn, CeFeTiNi, FeCrNi
23 Novartis Vaccines and Diagnostics Agrippal S1 no SiAlK, Si, SiMgFe, CaSiAl, SBaZn
24 Novartis vaccines Agrippal no Cr, Ca, SiCaAl, ZrSi, SBa, CuZn, SCa
25 Novartis Vaccines and Diagnostics S Fluad no CaSiAl, FeSiTi,SiMgAlFe, SBa
26 Novartis Vaccines and Diagnostics Menveo no CaSiAl, SiAlFe, FeCrNi, Fe, Al, SBa
27 Pfizer Prenevar 13 no FeCr
W, CaAlSi, Al, CaSiAlFe, FeS, FeCr, FeCrNi, Fe, , CaP, FeTiMn,
28 Pfizer Prevenar 13 no
Ba, SiMgAlFe
29 Pfizer Meningitec - ctrl no Cr, Si
30 Pfizer Meningitec - ctrl no FeCrNi, W
31 Pfizer Meningitec no CaSiAl, CaSi, SiAlFeTi, FeCrNi, W, Fe, Pb
32 Pfizer Meningitec no Cr (precipitates), Ca, AlSi
33 Pfizer Meningitec no W, SiCa, CaSi, Pb, FeCrNi, Cr
34 Wyeth Pharmaceutical - UK Meningitec no SiAlFe, SiAlTi, SiMgFe, W, Fe, Zr, Pb, Ca, Zn, FeCrNi
35 Sanofi Pasteur MSD-France Vaxigrip no Fe, FeCrNi, SiAlFe, AlSi, SiAlFeCr
36 Sanofi Pasteur MSD Stamaril Pasteur no CaSiAl, AlSi, Fe, SiMgFe, SiMgAlFe, CrSiFeCr, CrSiCuFe
37 Sanofi Pasteur MSD Gardasil AlPO4. 2H2O AlCuFe, PbBi, Pb, Bi, Fe
38 Sanifi Pasteur MSD Gardasil AlPO4. 2H2O CaAlSi, AlSi, SiMgFe, Al,Fe, AlCuFe, FeSiAl, BiBaS, Ti, TiAlSi
39 Sanofi Pasteur Vaxigrip no Ca, CrFe, FeCrNi, CaSZn, CaSiAlTiFe, Ag, Fe
40 Sanofi Pasteur Vaxigrip no SiMgFe, CaSiAl, AlSiFe, AlSi,FeCr, FeZn, Fe
41 Sanofi Pasteur MSD Repevax AlPO4 .2H2O Bi, Fe, AlSiFe, SiMg, SBa, Ca
Ti, Br, AuCuZn, Ca, SiZn, SiAuAgCu, SiMgFe,FeCrNi.
42 Sanofi Pasteur MSD S Repevax AlPO4.2H2O
AlSiMgTiMnCrFe, SiFeCrNi, FeAl
Si, SiFeCrNi, FeCrNi,FeNi, Fe, SCa, AlSiCa, CaAlSiFeV, SBa, Pt,
43 Sanofi Pasteur MSD M-M-R vaxPro no
PtAgBiFeCr
44 Virbac S.A. - Carros - France Feligen CRP no Ca,SiAl
Citation: Gatti AM, Montanari S. New quality-control investigations on vaccines: micro- and nanocontamination. Int J Vaccines Vaccin. 2017;4(1):7‒14.
DOI: 10.15406/ijvv.2017.04.00072
Copyright:
New quality-control investigations on vaccines: micro- and nanocontamination ©2017 Gatti et al. 10
Table 3 List of the debris’ number identified in each vaccine as single particle, clusters and aggregates. Characterization is made by shape, size range and
variability of the number of particles counted in each aggregate [in brackets]
Size Range Cluster Size Range Aggregate n.(Range Size
Name Total Debris n.
in mm n. in mm of Particles) Range mm
Allergoid NaCl precipitates / / / / /
Typhim Vi 394 0,1-2,5 3[9-350] Feb-35
Tetabulin 519 0,1-15 3[100-180] 25-60
Vivotif Berna 4 1,5-15
Inflexal V 103 0,1-17 1 20 3[35-45] 25-Oct
Anatetall 2 3-Jan
Morupar / / /
Mencevax ACWY 13 0,2-5
Infanrix 3 5-Jan 1 25
Infanrix hexa 1821 0,1-15 15[1820] 20-140
Infanrix hexa 162 0,3-7 12 60 2[7 debris] 3.5-44
Typherix 8 0,2-8 1 15
Priorix 641 0,05-30 1 10 3[600] 20-70
Engerix-B precipitates /
Varilrix 2723 0,1-4 36 [120-2000] 15-40
Fluarix 1317 0,1-40 3[83] 30-Jul
Cervarix 1569 0,2-3 2 10-May 4[1400] 30-Aug
Anatetall 47 0,05-40
Dif-Tet-All 111 0,2-3
Menjugate 73 0,1-5
Focetria 35 0,7-10
Agrippal S1 430 0,2-6 13 0.2-80 5[410] 20
Agrippal S1 1029 0,1-12 9[1025] 35-80
Agrippal 480 0,1-6 11[ 460] Feb-80
Fluad 605 0,2-15 4 25-Dec 6[ 600] 70
Menveo 452 0,1-13 4[430 ] 30-110
Prenevar 13 precipitates + 5 debris 7-Jan
Prevenar 13 precipitates + 81 debris 0,2-50 3 May-40 1 [60] 25
Meningitec 3 20-Oct
Meningitec 24 Aug-60
Meningitec 673 0,1-20 1 7 9[624] 5-110
Meningitec precipitates + 40 0,1-3,5 2[40] 25-70
Meningitec 177 0,2-10 3[165] 15-100
Meningitec 241 0,1-15 1 50 2[230] 50
Vaxigrip 86 0,1-7 2[50] 2-2,5
Stamaril Pasteur 152 0,1-7 2 7-May 3[145] 20-Apr
Gardasil 304 0,05-3 1[300] 15
Gardasil 454 0.1-30 2 20-Jul 9[445] May-60
Vaxigrip 304 0,1-10 1 13 2[300] 35
Vaxigrip 674 0,3-25 2 12-Feb 10[660] 9-150
Repevax 137 0,1-20 2[130] 40-50
Repevax 214 0,1-10 6[150] 30-May
M-M-R vaxPro 93 0,1-15 2[50] 15-Oct
Feligen CRP 92 0,1-12 1 12 1 (40 debris) 25
The study was aimed at verifying a possible physical covered with an adhesive carbon disc. The sample is immediately put
contamination. To do that, we performed a new kind of investigation inside a clean box in order to avoid any contamination and the box is
based on observations under a Field Emission Gun Environmental re-opened only for the sample to be inserted inside the FEG-ESEM
Electron Scanning Microscope (FEG-ESEM, Quanta 200, FEI, The chamber. We selected that particular type of microscope as it allows
Netherlands) equipped with the X-ray microprobe of an Energy to analyse watery and oily samples in low vacuum (from 10 to 130 Pa)
Dispersive Spectroscope (EDS, EDAX, Mahwah, NJ, USA) to detect at a high sensitivity.
the possible presence of inorganic, particulate contaminants and
When the water and saline the vaccine contains are evaporated,
identify their chemical composition.
the biological/physical components emerge on the filter and it is then
A drop of about 20microliter of vaccine is released from the possible to observe them. This type of microscope (low-vacuum
syringe on a 25-mm-diameter cellulose filter (Millipore, USA), inside observations) prevents the possible sample contamination and the
a flow cabinet. The filter is then deposited on an Aluminum stub creation of artefacts. The observations are made with different sensors
Citation: Gatti AM, Montanari S. New quality-control investigations on vaccines: micro- and nanocontamination. Int J Vaccines Vaccin. 2017;4(1):7‒14.
DOI: 10.15406/ijvv.2017.04.00072
Copyright:
New quality-control investigations on vaccines: micro- and nanocontamination ©2017 Gatti et al. 11
(SE: secondary-electron sensor and BSE: backscattered-electron Single inorganic particles or organic-inorganic aggregates are
sensor), and are performed at a pressure of 8.9 e-1 mbar, at energies identified, evaluated and counted. The counting procedure is repeated
ranging from 10 to 30kV to detect the particulate matter’s size, three times by three different operators, with an error lower than 10%.
morphology and its elemental composition. The method identifies When a layer of salts (Sodium chloride or Aluminum) is detected, we
clearly inorganic bodies with a higher atomic density (looking whiter) record the situation but we do not do body count.
than the biological substrate. So, organic entities are visible and easy
Results
to distinguish from inorganic ones. The method cannot distinguish
between proteins and organic adjuvants (e.g. squalene, glutamate,
The investigations verified the physical-chemical composition
proteins, etc.) or viruses, bacteria, bacteria’s DNA, endo-toxins and
of the vaccines considered according to the inorganic component as
bacteria’s waste, but their comparatively low atomic density allows
declared by the Producer. In detail, we verified the presence of saline
us to identify these entities as organic matter. In some vaccines, the
and Aluminum salts, but further presence of micro-, sub-micro- and
organic matter contains white-looking debris named aggregates, while
nanosized, inorganic, foreign bodies (ranging from 100nm to about
a high concentration or inorganic debris is called a cluster.
ten microns) was identified in all cases, whose presence was not
declared in the leaflets delivered in the package of the product (Table
2).Play Video
Figure 1a shows a layer of crystals of Sodium chloride (NaCl)
embedding salts of Aluminum phosphate (AlPO) in a drop of
4
Gardasil (anti-HPV vaccine by Merck) as the EDS spectrum (Figure
1b) shows. Saline is the fluid base to any vaccine preparation and
Aluminum salts or Aluminum hydroxide Al(OH)3 are the adjuvants
which are usually added.
Figure 1 Crystals of saline solution and Aluminum Phosphate and Looking at the area outside these precipitates but inside the liquid
corresponding EDS spectra. drop, we identified other things: single particles, clusters of particles
and aggregates (organic-inorganic composites) that are due to an
interaction of the inorganic particulate matter with the organic part
of the vaccine.
Figure 2a-2f shows the different typology of entities identified
in the vaccines (Repevax, Prevenar and Gardasil); single particles,
cluster of micro- and nanoparticles (<100nm) and aggregates with
their EDS spectra (Figure 2d-2f). The images (Figure 2a & 2d) show
debris of Aluminum, Silicon, Magnesium and Titanium; of Iron,
Chromium, Silicon and Calcium particles (Figure 2b & 2e) arranged
in a cluster, and Aluminum -Copper debris (Figure 2c & 2f) in an
aggregate.
Figure 2 Images of single particles, cluster of micro- and nanoparticles
(<100nm) andaggregates with their EDS spectra. They are respectively As can be seen, the particles are surrounded and embedded in a
composed of (a,b) Aluminum, Silicon, Magnesium, Titanium, Chromium, biological substrate. In all the samples analyzed, we identified particles
Manganese, Iron, (c,d) Iron, Silicon, Calcium Titanium, Chromium, (e,f)
containing: Lead (Typhym, Cervarix, Agrippal S1, Meningitec,
Aluminum, Copper. The arrows show the points where EDS spectra were
Gardasil) or stainless steel (Mencevax, Infarix Hexa, Cervarix.
taken.
Anatetall, Focetria, Agrippal S1, Menveo, Prevenar 13, Meningitec,
Vaxigrip, Stamaril Pasteur, Repevax and MMRvaxPro).
Figure 3a-3d show particles of Tungsten identified in drops of
Prevenar and Infarix (Aluminum, Tungsten, Calcium chloride).
Figure 4a-4d present singular debris found in Repevax (Silicon,
Gold, Silver) and Gardasil (Zirconium).
Some metallic particles made of Tungsten or stainless steel were
also identified. Other particles containing Zirconium, Hafnium,
Strontium and Aluminum (Vivotif, Meningetec); Tungsten, Nickel,
Iron (Priorix, Meningetec); Antimony (Menjugate kit); Chromium
(Meningetec); Gold or Gold, Zinc (Infarix Hexa, Repevax), or
Platinum, Silver, Bismuth, Iron, Chromium (MMRvaxPro) or
Lead,Bismuth (Gardasil) or Cerium (Agrippal S1) were also found.
The only Tungsten appears in 8/44 vaccines, while Chromium
(alone or in alloy with Iron and Nickel) in 25/44. The investigations
revealed that some particles are embedded in a biological substrate,
probably proteins, endo-toxins and residues of bacteria. As soon as a
particle comes in contact with proteic fluids, a nano-bio-interaction6
Figure 3 Images of Tungsten particles identified in drops of Prevenar and
Infarix. They are composed respectively of Tungsten, Aluminum, Iron but in occurs and a “protein corona” is formed.7‒10 The nano-bio-interaction
different concentrations. The arrows show the points where EDS spectra generates a bigger-sized compound that is not biodegradable and can
were taken. induce adverse effects, since it is not recognized as self by the body.
Citation: Gatti AM, Montanari S. New quality-control investigations on vaccines: micro- and nanocontamination. Int J Vaccines Vaccin. 2017;4(1):7‒14.
DOI: 10.15406/ijvv.2017.04.00072
Copyright:
New quality-control investigations on vaccines: micro- and nanocontamination ©2017 Gatti et al. 12
Nickel. (Figure 7a & 7b) present an area of Repevax where the
morphology of red cells - we cannot tell whether they are human or
animal- is clearly visible.
Figure 6 show an organic aggregate containing a debris made of Cerium, Iron,
Nickel, Titanium. The red arrow indicates the organic layer (less atomically
dense) that covers the Cerium particle.
Figure 4 Images show examples of nano biointeraction. The aggregate (a,b)
identified in Gardasil contains nanoparticles of Chlorine, Silicon, Aluminum,
Zirconium, while the debris found in Repevax contains Silicon, Gold, Silver
(c,d). The arrows show the points where EDS spectra were taken.
Figure 7 Image of an area in a Repevax drop where the morphology of red
cells (red arrows) were identified. It is impossible to know whether they are
human or animal origin. Among the debris of saline and Aluminum phosphate,
there is the presence of debris (white arrows) composed of Aluminum,
Bromine, Silicon, Potassium, Titanium.
Figure 5 show particles surrounded by an organic compound. They are
composed of Lead (a,b), Iron, Chromium, Nickel (stainless steel; c,d), Copper, Figure 8 Graph of the debris’ quantities identified in a 20 microl drop of
Tin, Lead (e,f). The arrows show the points where EDS spectra were taken. every vaccine.
Figure 5a-5f show examples of these nano-bio-interactions. Table 3 summarizes the number and morphology of the debris
Aggregates can be seen (stable composite entities) containing particles identified, in term of single particles, clusters of particles or
of Lead in Meningitec, (Figure 5a & 5b) of stainless steel (Iron, aggregates (organic-inorganic compounds), while Figure 8 shows the
Chromium and Nickel, Figure 5c & 5d) and of Copper, Zinc and Lead graph obtained calculating the total number of particles (particles plus
in Cervarix (Figure 5e & 5f). Similar aggregates, though in different clusters plus aggregates) identified for 20microl of every vaccine.
situations (patients suffering from leukemia or cryoglobulinemia),
Similar aggregates were already described by other scientists who
have already been described in literature.
identified them in the blood e.g. in leukemic patients15 and in subjects
The link between these two entities generates an unfolding of the affected by cryoglobulinemia.16
proteins that can induce an autoimmune effect once those proteins are
Not all the vaccines analyzed contain the same contamination,
injected into humans.
though the same vaccine belonging to different batches and, in
Figure 6a & 6b show one of the foreign bodies identified in some cases, coming from different countries can contain a similar
Agrippal. The particle is composed of Cerium, Iron, Titanium and contamination (e.g. the vaccines by Glaxo Infarix, Typherix and
Citation: Gatti AM, Montanari S. New quality-control investigations on vaccines: micro- and nanocontamination. Int J Vaccines Vaccin. 2017;4(1):7‒14.
DOI: 10.15406/ijvv.2017.04.00072
Copyright:
New quality-control investigations on vaccines: micro- and nanocontamination ©2017 Gatti et al. 13
Priorix contain Tungsten. Tungsten was also identified in Menjugate not be present in any injectable medicament, let alone in vaccines,
kit by Novartis, and Prevenar, Meningitec by Pfizer and Meningitec more in particular those meant for infants.
by Wyeth).
Other forms of so-far unknown contaminations have recently been
Feligen, the only veterinary vaccine tested, proved to be the only observed and, in any case, vaccines contain components that could
sample free from inorganic contamination, while Allergoid generates themselves be the cause of adverse effects. It is a well-known fact in
a layer of inorganic salts so thick that it does not allow to detect other toxicology that contaminants exert a mutual, synergic effect, and as
particulate contaminants. the number of contaminants increases, the effects grow less and less
predictable. The more so when some substances are unknown.
Discussion
As a matter of fact, no exhaustive and reliable official data exist
The quantity of foreign bodies detected and, in some cases, their on the side-effects induced by vaccines. The episodic evidence
unusual chemical compositions baffled us. The inorganic particles reported by people allegedly damaged by vaccines is twofold: some
identified are neither biocompatible nor biodegradable, that means that say the damage occurred and became visible within a few hours from
they are biopersistent and can induce effects that can become evident administration, and some maintain that it was a matter of some weeks.
either immediately close to injection time or after a certain time from Though we have no indisputable evidence as to the reliability of those
administration. It is important to remember that particles (crystals and attestations, we can put forward a hypothesis to explain the different
not molecules) are bodies foreign to the organism and they behave as phenomena. In the former case, the pollutants contained in the drug
such. More in particular, their toxicity is in some respects different have reached the brain and, depending on the anatomical site interested,
from that of the chemical elements composing them, adding to that have induced a reaction. If that is the case, the whole phenomenon
toxicity which, in any case, is still there, that typical of foreign bodies. is very rapid. In the latter circumstance, the pollutants reached the
For that reason, they induce an inflammatory reaction. microbiota, thus interfering with the production of enzymes necessary
to carry out neurological functions.32‒35 That possibility takes time,
After being injected, those microparticles, nanoparticles and
as it involves the production of chemical compounds in a sufficient
aggregates can stay around the injection site forming swellings and
quantity, and an elapse of some weeks between injection and clinical
granulomas.17 But they can also be carried by the blood circulation,
evidence is reasonable. Of course, ours is no more than a hypothesis
escaping any attempt to guess what will be their final destination.
open to discussion and in need of proof, hoping that a chance of
We believe that in many cases they get distributed throughout the
further investigation is allowed.
body without causing any visible reaction, but it is also likely that,
in some circumstances, they reach some organ, none excluded and Conclusion
including the microbiota, in a fair quantity. As happens with all
foreign bodies, particularly that small, they induce an inflammatory The analyses carried out show that in all samples checked vaccines
reaction that is chronic because most of those particles cannot be contain non biocompatible and bio-persistent foreign bodies which
degraded. Furthermore, the protein-corona effect (due to a nano- are not declared by the Producers, against which the body reacts in
bio-interaction18 can produce organic/inorganic composite particles any case. This new investigation represents a new quality control that
capable of stimulating the immune system in an undesirable way.19‒22 can be adopted to assess the safety of a vaccine. Our hypothesis is that
It is impossible not to add that particles the size often observed in this contamination is unintentional, since it is probably due to polluted
vaccines can enter cell nuclei and interact with the DNA.23 components or procedures of industrial processes (e.g. filtrations)
used to produce vaccines, not investigated and not detected by the
In some cases, e.g. as occurs with Iron and some Iron alloys, they
Producers. If our hypothesis is actually the case, a close inspection of
can corrode and the corrosion products exert a toxicity affecting the
the working places and the full knowledge of the whole procedure of
tissues.24‒26
vaccine preparation would probably allow to eliminate the problem.
The detection of presence of Aluminum and NaCl salts is obvious
A further purification of the vaccines could improve their quality
as they are substances used by the Producers and declared as
and could probably decrease the number and seriousness of the
components, but other materials are not supposed to be in the vaccine
adverse incidental effects.
or in any other injectable drug, at that, and, in any case, Aluminum has
already been linked with neurological diseases.27‒29 Acknowledgments
Given the contaminations we observed in all samples of human- The Authors are indebted to Dr. Federico Capitani, Dr. Laura
use vaccines, adverse effects after the injection of those vaccines are Valentini and Ms. Lavinia Nitu for their technical assistance. The
possible and credible and have the character of randomness, since opinions and conclusions are not necessarily those of the Institution.
they depend on where the contaminants are carried by the blood
Conflicts of interest
circulation. It is only obvious that similar quantities of these foreign
bodies can have a more serious impact on very small organisms like
Author declares there are no conflicts of interest.
those of children. Their presence in the muscles, due an extravasation
from the blood, could heavily impair the muscle functionality.30,31 Funding
We come across particles with chemical compositions, similar None.
to those found in the vaccines we analyzed, when we study cases of
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Citation: Gatti AM, Montanari S. New quality-control investigations on vaccines: micro- and nanocontamination. Int J Vaccines Vaccin. 2017;4(1):7‒14.
DOI: 10.15406/ijvv.2017.04.00072